Isolation of the encoding gene for a thermostable α-glucosidase from Geobacillus stearothermophilus strain RM and its expression in Escherichia coli

High temperature catalysis increases substrate solubility and carbohydrate hydrolysis, therefore investigating new thermostable alpha-glucosidase is an attractive option. A total of seven isolates were obtained from two different hot spring locations in Malaysia, namely Telaga Air Hangat Langkawi and Slim River. Preliminary study on four selected isolates revealed that production media 1 (PM 1) and production media 2 (PM 2) were the best media to support alpha-glucosidase production. The optimum growth temperatures were 50-70 degrees C while their optimum temperature for alpha-glucosidase production was 55 degrees C. Quantitative screening indicated that isolate L3 which was identified as Geobacillus stearothermophilus strain RM was the best alpha-glucosidase producer with 1.47 U/ml at 55 degrees C after 72 h. The production of alpha-glucosidase was also found to be growth associated up to stationary phase. Identification on the basis of morphological characteristics, biochemical studies and 16S rRNA analysis were carried out and 16S rRNA identification revealed that this isolate showed 99% similarity to G. stearothermophilus. The alpha-glucosidase gene had been successfully amplified from this identified bacterium via degenerate primer and the complete gene was cloned and expressed into Escherichia coli with 5 U/ml activity which was 5-folds higher compared to the wild type. As a conclusion, E. coli system successfully increased the yield of alpha-glucosidase production.